Tooth enamel is a highly mineralized hard tissue, uniquely comprised of millions of hexagonal carbonated hydroxyapatite (HAP) crystals. These crystals are very thin and extremely long, which determine excellent mechanical properties of tooth enamel. Mineralized enamel crystals develop from a layer of enamel protein matrix that is predominated by amelogenins (>90%), which forms special nanostructures to modulate crystal formation. Amelogenins are gradually and completely removed by enamel proteinases MMP20 and KLK4 to form highly mineralized enamel at the end of maturation stage. The interactions between crystal, amelogenin and proteinase dynamically and delicately controlled the growth rate, direction and morphology of enamel crystals. We hypothesize that the binding of amelogenin to apatite crystal changes the conformation of amelogenin adsorbed on crystals and results in preferential degradation of amelogenin from crystal surface by proteinases. The amelogenin adsorption and degradation are also specific on different crystal planes, which drive the crystal to elongate primarily along the c axis and widen/thicken along a and b axes in different developmental stages of amelogenesis. In addition, the N-terminal proline 70 is involved in the plane-specific adsorption and degradation of amelogenin. The mutation of the proline (P70T, linked to a type of amelogenesis imperfect) interferes with crystal- amelogenin-proteinase interactions, resulting in abnormal enamel mineralization and morphology. Three specific aims are proposed to evaluate the hypothesis: Specific Aim 1, to characterize and compare (001) and (hk0) planes of oriented HAP crystals; Specific Aim 2, to determine amelogenin-crystal interactions by identifying the binding domains, analyzing binding affinities and kinetics, and investigating the binding patterns of amelogenins on (001) and (hk0) planes of HAP crystals; Specific Aim 3, To investigate amelogenin-proteinases interactions before and after amelogenin binding to apatite crystals and determine how the interactions affect the growth of crystal. The amelogenin carrying P70T mutation will be also used to test the effects of the mutation on crystal- amelogenin-proteinase interactions. The proposed studies will integrate the roles of HAP, amelogenin and proteinase into an interactive unity, and study how the interactions modulate the enamel crystal growth and morphology. The findings collected in this study will build new concepts to advance our knowledge of the unique principle of amelogenin-mediated crystal growth and help us to better understand the fundamental mechanism of AI. The new concepts may be also useful for future acellular tooth enamel repair and regeneration.